title: Fluorescent probes for investigation of isoprenoid configuration and size discrimination by bactoprenol-utilizing enzymes.
abstract: Undecaprenyl Pyrophosphate Synthase (UPPS) is an enzyme critical to the production of complex polysaccharides in bacteria, as it produces the crucial bactoprenol scaffold on which these materials are assembled. Methods to characterize the systems associated with polysaccharide production are non-trivial, in part due to the lack of chemical tools to investigate their assembly. In this report, we develop a new fluorescent tool using UPPS to incorporate a powerful fluorescent anthranilamide moiety into bactoprenol. The activity of this analogue in polysaccharide biosynthesis is then tested with the initiating hexose-1-phosphate transferases involved in Capsular Polysaccharide A biosynthesis in the symbiont Bacteroides fragilis and the asparagine-linked glycosylation system of the pathogenic Campylobacter jejuni. In addition, it is shown that the UPPS used to make this probe is not specific for E-configured isoprenoid substrates and that elongation by UPPS is required for activity with the downstream enzymes.
authors.affiliation: University of North Carolina at Charlotte, Department of Chemistry, 9201 University City Blvd., Charlotte, NC 28223-0001, United States.
authors.forename: Anahita Z
authors.initials: AZ
authors.lastname: Mostafavi
authors.forename: Donovan K
authors.initials: DK
authors.lastname: Lujan
authors.forename: Katelyn M
authors.initials: KM
authors.lastname: Erickson
authors.forename: Christina D
authors.initials: CD
authors.lastname: Martinez
authors.forename: Jerry M
authors.initials: JM
authors.lastname: Troutman
mesh_terms: D019883:Alkyl and Aryl Transferases / Q000378:metabolism*; D001439:Bacteroides / Q000378:metabolism; D016123:Campylobacter jejuni / Q000201:enzymology / Q000378:metabolism; D005456:Fluorescent Dyes / Q000737:chemistry*; D006031:Glycosylation; D011134:Polysaccharides / Q000096:biosynthesis / Q000737:chemistry; D013237:Stereoisomerism; D013379:Substrate Specificity; D013729:Terpenes / Q000737:chemistry* / Q000378:metabolism; D062367:ortho-Aminobenzoates / Q000737:chemistry
chemical_list: D005456:Fluorescent Dyes; D011134:Polysaccharides; D013729:Terpenes; D062367:ortho-Aminobenzoates; C000146:bactoprenol; D019883:Alkyl and Aryl Transferases; C023266:undecaprenyl pyrophosphate synthetase; C000219:anthranilamide
keywords: Bactoprenol; Isoprenoid; UPPS; Undecaprenol; cis-Prenyltransferase